Acetonitrile-phosphate Buffer Research Articles

Potentiometry in determining the antioxidant capacity of compounds of different hydrophilicity in the joint presence using mixed solvents and surfactants.

In this work, for the first time, the potentiometric method was used to determine the antioxidant capacity (AOC) of compounds of different hydrophilicity in the joint presence using mixed solvents and surfactants. The AOC of model solutions of antioxidants of different hydrophilicity was determined separately and in the joint presence in the media of phosphate buffer-surfactant and mixed solvents-surfactant, using as an example the ascorbic acid and the α-tocopherol. It was shown that the surfactant Triton X-100 is able to solubilize α-tocopherol under the selected conditions, allows to obtain reproducible and accurate results, and has less effect on the equilibrium rate of the K3[Fe(CN)6]/K4[Fe(CN)6] system. Phosphate buffer-ethanol and phosphate buffer-acetonitrile media in the 3:2 ratio with Triton X-100 (5mmol/dm3) were chosen as the analysis conditions (RSD ≤ 6%). The range of determined concentrations was (0.006-0.5)m mmol/dm3 in phosphate buffer-ethanol and (0.006-0.3)m mmol/dm3 in phosphate buffer-acetonitrile. AOC of raw materials infusions was determined in selected media. The positive correlation with the well-known Folin-Ciocalteu assay was obtained.

Analytical Purity Determinations of Universal Food-Spice Curcuma longa through a QbD Validated HPLC Approach with Critical Parametric Predictors and Operable-Design’s Monte Carlo Simulations: Analysis of Extracts, Forced-Degradants, and Capsules and Tablets-Based Pharmaceutical Dosage Forms

Applications of analytical quality by design (QbD) approach for developing HPLC (High Performance Liquid Chromatography) methods for food components assays, and separations of complex natural product mixtures, are still limited. The current study developed and validated, for the first time, a stability-indicating HPLC method for simultaneous determinations of curcuminoids in Curcuma longa extracts, tablets, capsules, and curcuminoids' forced degradants under different experimental conditions. Towards separation strategy, critical method parameters (CMPs) were defined as the mobile phase solvents' percent-ratio, the pH of the mobile phase, and the stationary-phase column temperature, while the peaks resolution, retention time, and the number of theoretical plates were recognized as the critical method attributes (CMAs). Factorial experimental designs were used for method development, validation, and robustness evaluation of the procedure. The Monte Carlo simulation evaluated the developing method's operability, and that ensured the concurrent detections of curcuminoids in natural extracts, commercial-grade pharmaceutical dosage-forms, and the forced degradants of the curcuminoids in a single mixture. The optimum separations were accomplished using the mobile phase, consisting of an acetonitrile-phosphate buffer (54:46 v/v, 0.1 mM) with 1.0 mL/min flow rate, 33 °C column temperature, and 385 nm wavelength for UV (Ultra Violet) spectral detections. The method is specific, linear (R2 ≥ 0.999), precise (% RSD < 1.67%), and accurate (% recovery 98.76-99.89%), with LOD (Limit of Detection) and LOQ (Limit of Quantitation) at 0.024 and 0.075 µg/mL for the curcumin, 0.0105 µg/mL and 0.319 µg/mL for demethoxycurcumin, and 0.335 µg/mL and 1.015 µg/mL for the bisdemethoxycurcumin, respectively. The method is compatible, robust, precise, reproducible, and accurately quantifies the composition of the analyte mixture. It exemplifies the use of the QbD approach in acquiring design details for developing an improved analytical detection and quantification method.

METHOD DEVELOPMENT AND VALIDATION OF CEFOPERAZONE AND SULBACTAM IN DRIED BLOOD SPOTS BY HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY PHOTODIODE ARRAY DETECTOR

Objective: The primary purpose of this research was to develop a simple, precise, fast, and accurate method for measuring cefoperazone and sulbactam simultaneously in dried blood spots (DBS) using HPLC PDA. Methods: A simplified analytical method for quantifying cefoperazone and sulbactam in DBS samples using a High-Performance Liquid Chromatography photodiode array detector with isocratic elution was developed and validated. The best chromatographic conditions were obtained by using a reversed-phase column (250 x 4.6 mm; 5 mm); phosphate buffer 10 mmol pH 3.2–acetonitrile (83:17, v/v) as a mobile phase; a flow rate of 1.0 ml/min; a column temperature of 35 °C; a photodiode array detector at 210 nm, and cefuroxime as internal standard. Samples were prepared by liquid-liquid extraction with 100 ml hydrochloric acid 0.5 mol/l and 1000 ml ethyl acetate, evaporated with nitrogen and reconstituted with 100 mL phosphate buffer–acetonitrile (4:1). Results: The total chromatography run time was 15 min, and the elution times for sulbactam, cefoperazone, and IS (cefuroxime) were 3.46, 10.221, and 6.987 min, respectively. A linear response function was established at 0.5-30 mg/ml with (r) 0.995 for sulbactam and 2.5-250 mg/ml with (r) 0.999 for cefoperazone in dried blood spots. The lower limit quantification (LLOQ) concentration of sulbactam 1 mg/ml and cefoperazone were 5 mg/ml. Conclusion: This method has successfully fulfilled the validation requirement referring to the 2011 EMA and 2018 FDA guidelines.

Green Liquid Chromatography Method for the Determination of Related Substances Present in Olopatadine HCl Nasal Spray Formulation, Robustness by Design Expert.

Dual therapeutic nature drug mast cell stabilizer and histamine receptor antagonist olopatadine hydrochloride (OPT) nasal spray does not have an official monograph, and no literature is available. Eye drops formulation had the official monograph for impurities, but the determination was done in two methods. A simple and effective green liquid chromatography method to develop and validate for the related substances of OPT nasal spray formulation. A 25 min gradient method was employed to separate impurities and OPT with a 1.0 mL/min flow rate using a Boston green C8 (150 mm × 4.6 mm, 5 µm) HPLC column. The set wavelength and column oven temperatures were 299 nm and 30°C, respectively. pH 3.5 phosphate buffer-acetonitrile in the ratio of (70:30, v/v) as mobile phase A and (50:50, v/v) ratio as mobile phase B. A Quality by Design (QbD) based Design of Experiments (DoE) was employed to evaluate the robustness characteristics of the analytical method validation. The obtained RSD from the precision and intermediate precision was 0.4 to 4.1%. The % recovery of the impurities from LOQ to 150% of specification level was 87.5 to 110.3%. The linear regression curves for the impurities with a correlation coefficient of >0.999 indicate that all peak responses are linear with the concentration. The sample and standard solutions were stable for 24 h at benchtop and refrigerator conditions. All the critical peaks were well separated from the forced degradation studies' diluent, placebo, and generated degradation peaks. The method validation data and QbD based robustness study results indicate that the developed impurities method fits the routine quality control laboratory use. National Environmental Index (NMEI), Green Analytical Procedure Index (GAPI), Analytical Eco-scale and Analytical Greenness (AGREE) tools expressed the method's greenness. The proposed method is QbD utilized and green chemistry assessed impurities determination method for OPT in nasal spray formulation.

Development and Validation of RP-HPLC Method for the Determination of Dexrabeprazole Sodium

A rapid and simple liquid chromatographic method was developed for the quantitative determination of dexrabeprazole sodium in the tablet dosage form. The reverse-phase chromatographic analysis was carried out by using a C-18 column. The acetonitrile-phosphate buffer mixture was used as the mobile phase. The obtained retention time was 4.33∓0.02 min. by using 50:50 (v:v) acetonitrile:phosphate buffer(pH 7) and 1.0 mL/min. flow rate. Quantification of the analyte was based on measuring the peaks areas at 272 nm. The analytical performance of the developed RP-HPLC method was validated with respect to accuracy, precision, linearity, stability, and robustness. The obtained linearity range was 77-143 mg/L and the correlation coefficient was 0.9989. The obtained LOD and LOQ values were 0.010 mg/L and 0.034 mg/L, respectively.

Development of a Validated Stability-Indicating HPLC Method for Vinpocetine in the Presence of Potential Impurities in Tablet Dosage Form by Multiresponse Optimization.

Vinpocetine has been prescribed for the treatment of ischemic brain diseases for many years. The drug, which has side effects such as headache, flushing, and decreased blood pressure, is not found in nature, but it can be synthesized by several approaches. A simple, rapid, selective, stability-indicating high-performance liquid chromatographic (HPLC) method was developed for the simultaneous estimation of vinpocetine and their potential impurities in a tablet formulation. Optimum HPLC conditions were tried to be determined by a statistical experimental design method. The proposed method was validated as per the ICH guidelines. Stress study was used to demonstrate the stability-indicating ability of the developed method in the quantification of vinpocetine and potential impurities within the same run. According to multiresponse optimization using the Derringer's desirability function, the mobile phase consisted of an acetonitrile-phosphate buffer (pH 6.0) in the ratio of 65:35 (v/v) at a flow rate of 1.7 mL/min. Significant degradations were observed for the drug product under acid hydrolysis and alkali hydrolysis. The new method showed reasonable detection and quantification limits with good selectivity, precision, linearity, and recovery. These validation results have shown that this method is suitable for quantitative determination of vinpocetine and its impurities in quality control laboratories. A reliable and rapid HPLC method optimized with response surface methodology and multiresponse optimization based on Derringer's desirability function was developed for the simultaneous analysis of vinpocetine and its impurities in a tablet formulation.

Determination of residual dimethylaminopropylamine in aminopropylamides of fatty acid using high-performance liquid chromatography

A procedure for determination of the residual content of N,N-dimethyl-1,3-propanediamine (DMAPA) during synthesis of N,N-dimethylaminopropylamides of fatty acids (DMAPA FA) from coconut oil has been developed. The analysis was performed using high performance liquid chromatography on a Shimadzu LC-20 Prominence device equipped with a diode array detector and reversed phase column GL Sciences Inertsil ODS-3. Precolumn derivatization was carried out with a 13% — dansyl chloride (5-(dimethylamino)naphthalene-1-sulfonyl chloride) solution in acetone to increase the analyte response. The selected composition of the mobile phase — acetonitrile and triethylamine phosphate buffer solution (pH 3.0) in a volume ratio of 2:3 and optimized chromatographic conditions provided a clear peak of DMAPA with a retention time of 8.7 – 8.9 min. The proposed method provides determination of 0.02 – 10% DMAPA in DMAPAFA samples. Correctness of the procedure was confirmed in spike tests. The results obtained can be used for assessing the degree of conversion of starting materials in the synthesis of N,N-dimethylaminopropylamides of fatty acids, as well as for forecasting the content of N,N-dimethyl-1,3-propanediamine in the products on their base.

A Comparative Estimation of Alprazolam in Pharmaceutical Formulations by Validated HPLC and HPTLC Techniques

The aim of present work is to develop and validate two simple and specific methods for the estimation of alprazolam hydrochloride (a psychoactive drug) in pharmaceutical formulations. The first method involves HPLC separation on the Cosmosil C-8 column using acetonitrile and potassium phosphate buffer (pH 6.0±0.1) (40:60 v/v) as mobile phase. The peak was obtained at 3.01±0.02 min. The effluent was monitored at 254 nm. The second method is based on HPTLC using Acetic acid: water: methanol: ethyl acetate (2:15:20:80 v/v/v/ v) as mobile phase and detection of the spot was carried out at 254 nm. The Rf value was found to be 0.76±0.02. The methods were validated as per ICH guidelines. The linearity curves were found to be linear over the concentration range 50-150 μg/ ml for HPLC and 50-400 ng/spot for HPTLC with a high correlation coefficient (R2 = 0.991 and 0.9987 respectively). The limit of detection and limit of quantification were found to be 1.66 and 5.04 μg/ml for HPLC and14.84 and 44.097 μg/ ml for HPTLC, respectively. The proposed methods are successfully used for the quality control of alprazolam in commercial form. Statistical analysis results confirmed that the methods are selective and precise.

Rapid and Sensitive Liquid Chromatographic Method for Determination of Anastrozole in Different Polymer–Lipid Hybrid Nanoparticles

Anastrozole, an aromatase inhibitor drug, is used for the treatment of breast cancer in pre- and postmenopausal women. Anastrozole's incorporation into nanoparticulate carriers would enhance its therapeutic performance. To perceive the exact loaded amount of drug in nanocarriers, a valid analytical method is required. The reversed-phase high-performance liquid chromatography (RP-HPLC) method was developed and validated by using the C18 column, 150 × 4.6 mm, 5 µm particle size, in isocratic mobile phase composed of 50:50 V/V (volume/volume) acetonitrile-phosphate buffer (pH 3) flowing at a rate of 1.0 mL/min, and a diode array detector (DAD) set at λmax = 215 nm. The validation parameters such as linearity, accuracy, specificity, precision, and robustness have proven the accuracy of the method, with the relative standard deviation percentage (% RSD) values < 2. The limit of detection of the method was found equal to 0.0150 µg/mL, and the limit of quantitation was 0.0607 µg/mL. The percent recovery of sample was in the range of 98.04-99.25%. The method has the advantage of being rapid with a drug retention time of 2.767 min, specific in terms of resolution of peaks void of interference with any of the excipients, and high reproducibility. This makes it highly applicable for quality control purposes.

Pharmacokinetic Study and Incurred Sample Stability of Esomeprazole in Dried Blood Spot Sample Using High Performance Liquid Chromatography-Photodiode Array

In the past years, Esomeprazole (EMP) was analyzed in human plasma samples, which still has stability issues; thus, the new biosampling technique known as Dried Blood Spot (DBS) might solve the issue. This research aims to evaluate the incurred sample stability of esomeprazole in dried blood spot using high performance liquid chromatography-photodiode array with lansoprazole as an internal standard. The analytical separation was performed on a C-18 column (Waters, Sunfire™ 5 μm; 250 × 4.6 mm) at 40 °C. The mobile phase used was acetonitrile–phosphate buffer pH 7.6 (40:60% v/v) with a flow rate of 1.00 mL/min; and was detected at 300 nm. The analyte was extracted from dried blood spot by methanol. Incurred sample stability was evaluated from 6 healthy subjects on day 0, 7, 14, and 28, respectively. This method was linear in the range concentration of 70–1400 ng/mL with r &gt; 0.98. Pharmacokinetic study shows that the average of AUC0–t of EMP in the DBS sample was 1765.41 ngh/mL. The highest percent difference value of esomeprazole’s incurred samples stability on day 7, 14, and 28 from 6 healthy subjects were 9.81%. This result fulfilled the acceptance criteria, which is the percent difference should not be greater than 20%, and 67% of total samples have to fulfill the criteria. The incurred sample stability result showed that esomeprazole was stable in the DBS sample at least until 28 days with the highest value of percent difference is 9.81%.

Comparative study of electrospray mass spectrometry and first derivative method and validation by HPLC method

Three different methods are described for the simultaneous determination of amoxicillin trihydrate (AMX) and potassium klavulanate (KLV) in synthetic mixtures and pharmaceutical tablets. The first method depends on the first-derivative ultraviolet spectrophotometry with zero-crossing measurement. The second method is based on mass spectrometry utilizing electropspray ionization. For the electrospray studies in negative and positive ion modes, aspirin (ASP) and clindamycin (CLN) were used, respectively as internal standards for quantification. The calibration function was established in the same concentration ranges as in the first derivative method. A reversed-phase high performance liquid chromatographic (RP-HPLC) method involving ultraviolet detection (γ = 220 nm) was developed for the analysis of AMX and KLV. Chromatography was carried out on a C-18 column with mobile phase comprising of phosphate buffer-acetonitrile (40:60, v/v). The proposed methods have been validated with regard to the selectivity, detection limit, recovery, accuracy and precision. For both drugs, methods were found to be selective, linear (R ≈ 0.99), accurate (recovery = 100-105%) and precise (<3% RSD) in the range of 21-49 μg/mL for AMX and 3-7 μg/mL for KLV. The limit-of-detection and limit-of-quantification of the method were determined for three methods.

Determination of 13 Vitamin B and the Related Compounds Using HPLC with UV Detection and Application to Food Supplements

A high-performance liquid chromatographic method was developed for the separation and determination of 13 vitamin B compounds, including thiamin, riboflavin, riboflavin 5′-monophosphate, pyridoxine, hydroxocobalamin, cyanocobalamin, adenosylcobalamin, methylcobalamin, niacin, niacinamide, pantothenic acid, folic acid, and biotin. The method consisted of an ODS column, a gradient elution using pH 3.0 phosphate buffer–acetonitrile, ion-pairing reagent as mobile phase at 1.0 mL min−1, and UV detection at 210 nm. As a result, the 13 vitamin B compounds were separated in 60 min, and the specificity, linearity, range, limit of detection, limit of quantitation, intra- and inter-day precision, and recovery were satisfactory. The limit of detections ranged from 0.03 to 0.41 µg mL−1. Intra- and inter-day precision of peak area, expressed as RSD, were 0.542–5.144% and 0.216–6.367%, respectively. The method was used to evaluate vitamin B compounds in energy drinks and multivitamin pills. In case of energy drink analysis, the method identified and evaluated not only vitamin B compounds but also major ingredients, such as caffeine, vanillin, and benzoic acid. The vitamin B compounds in energy drinks and multivitamin pills were also quantitated and compared with values listed on each product label. Each quantitative value was close to the label values, suggesting high quantitative ability of the developed method.

تحليل المورفين، المورفين -3 غلوكورونيد، المورفين -6-غلوكورونيد، الكودايين والكودايين -6-الغلوكورونيد باستخدام جهاز الفصل السائل عالي الكفاءة

في علم السموم الشرعي والتفسير السريري، هناك حاجة كبيرة لتحديد واكتشاف العقاقير الأفيونية ومستقلبات الجلوكورونيد المختلفة. عادة ما يتم تحديد الجلوكورونيدات عن طريق انشقاق رابطة الجلوكورونيد مع إنزيم (على سبيل المثال، جلوكورونيداز) أو التحلل المائي الحمضي لإنتاج المركب الأصلي، الذي يتم اكتشافه لاحقًا. بالنسبة لتحليل الطيف الكتلي للغاز GC-MS، قد يكون هذا عن طريق الاشتقاق إلى شكل أكثر تقلبًا أو ثباتًا. الكشف المباشر عن اتحادات الجلوكورونيد يتغلب على القيود الحرجة لهذه الطرق. في هذا البحث، اختبار كروماتوجرافي السائل عالي الأداء وعكسي الطور عكسي انتقائي مع شطف التدرج والكشف عن الأشعة فوق البنفسجية للمورفين، المورفين 3-الجلوكورونيد، المورفين 6-الجلوكورونيد، الكوكايين، الكودايين -6-الجلوكورونيد والهيدروكودون تم تطويره. تم إجراء الفصل على عمود تحليلي C18 (سلسلة الكربون من Octadecyl) (150  2.0 مم، 5 ميكرون) واكتشفه كاشف الأشعة فوق البنفسجية (UV) عند 210 نانومتر. يتكون الطور المتحرك من مخزن مؤقت من الأسيتون تريل - الفوسفات (0.0125 ، درجة الحموضة 7.5) مع شطف في وضع التدرج تتراوح من 7.5 إلى 60% من الأسيتون تريل على مدى 25 دقيقة (دقيقة) مع معدل تدفق 0.2 مل / دقيقة. كانت منحنيات المعايرة خطية (نطاق R2 من 0.997 إلى 0.999) في نطاق التركيز 0.025-2 ميكروغرام / مل لكل التحليلات، باستخدام الهيدروكودون كمعيار داخلي. لذلك يهدف هذا السلوك إلى تطوير طريقة فعالة لفصل المورفين والكوديين والغلوكورونيدات بواسطة HPLC-UV وتقييم استخدام نصائح SPME LCلاستخراج المورفين والكوديين والجلوكورونيدات من عينات البول.

Determination of free clindamycin, flucloxacillin or tedizolid in plasma: Pay attention to physiological conditions when using ultrafiltration.

Pharmacokinetic/pharmacodynamic indices of anti-infective drugs should be referenced to free drug concentrations. In the present study, clindamycin, flucloxacillin and tedizolid have been determined in human plasma by HPLC-UV. The drugs were separated isocratically within 3-6 min on a C18 column using mixtures of phosphate buffer-acetonitrile of pH 7.1-7.2. Sample treatment for the determination of total drug concentrations in plasma included extraction/back-extraction (clindamycin) or protein precipitation (flucloxacillin, tedizolid). The free drug concentrations were determined after ultrafiltration. An ultrafiltration device with a membrane consisting of regenerated cellulose proved to be suitable for all drugs. Maintaining a physiological pH was crucial for clindamycin, whereas maintaining body temperature was essential for tedizolid. The methods were applied to the analysis of total and free drug concentrations in clinical samples and were sufficiently sensitive for pharmacokinetic studies and therapeutic drug monitoring.

THE OPTIMIZATION OF RP-HPLC CONDITION USING RESPONSE SURFACE METHODOLOGY BOX-BEHNKEN DESIGN FOR SIMULTANEOUS DETERMINATION OF METFORMIN HCL AND GLIMEPIRIDE IN SPIKED PLASMA

Objective: Aim of this study was to develop and validate the RP-HPLC method using Box-Behnken Design (BBD) for simultaneous analysis metformin HCl and glimepiride in spiked plasma.&#x0D; Methods: The chromatographic system was comprised of acetonitrile-phosphate buffer 0.0125 M+Sodium Dodecyl Sulphate (SDS) 1 mmol as a mobile phase and Ascentis® Phenyl C18 (250 x 4.6 mm i.d.; 5 µm) column as a stationary phase with UV detector at 210 nm. Three independent variables included phosphate buffer (%), pH and flow rate were optimized using Box-Behnken Design. The observed responses were retention time, peak area and resolution.&#x0D; Results: The predicted optimum condition of the RP-HPLC system consisted of phosphate buffer solution of 72%, pH at 4.3 and flow rate at 0.8 ml/min. By using this condition, the duration of analysis was more than 18 min, so it was necessary to modify the flow rate to be 1.0 ml/min to get shorter analysis duration. This condition was then applied to analyze metformin and glimepiride in spiked plasma and validated according to the EMA guideline. AUC of interfering components at the IS retention time between 588-1092 mV, the linearity of metformin was 0.9993 and glimepiride was 0.9991, accuracy and precision were between-13.33% until 16.08%, dilution integrity and metformin stability studies were between-4.01% until 11.82%, and for glimepiride stability studies were between-37.48% until-4.76%.&#x0D; Conclusion: Box-Behnken Design can help optimize the HPLC system, and the optimum condition was valid to analyze metformin and glimepiride in spiked plasma by considering the storage time of plasma samples.

Determinación y comparación cualitativa de cannabinoides presentes en productos naturales comerciales del departamento del Cauca-Colombia

Actualmente, hay un creciente interés por el estudio de Cannabis sativa y sus componentes ya que se le atribuye propiedades terapéuticas en el tratamiento de enfermedades. En Colombia y específicamente en el departamento del Cauca se comercializan productos de cannabis tanto para fines no medicinales como terapéuticos. En consecuencia, es necesario el análisis de estos productos de manera que se pueda conocer la composición de los mismos y el posible efecto que pueda tener sobre la salud.El análisis de los componentes de estos productos se llevó a cabo empleando la cromatografía líquida de alta resolución (CLAR) y espectrometría de masas (EM), de tal manera que permitieron la identificación de las principales especies cannabinoides; Δ9tetrahidrocannabinol (THC), cannabidiol (CBD), cannabinol (CBN), cannabigerol (CBG). La separación de los analitos se llevó a cabo mediante la implementación de una columna analítica C18 de fase reversa, elución isocrática 1 mL/min, presión del sistema 800 PSI, una mezcla de acetonitrilo ACN y buffer fosfato (KHPO4) en relación 65/35 como fase móvil, volumen de inyección de 10 µL, un tiempo de análisis de 15 min, y detección a 220 nm.

A reliable, simple and cost-efficient TLC-HPLC method for simultaneously determining florfenicol and florfenicol amine in porcine urine: application to residue surveillance

ABSTRACT Violative residues of florfenicol (FF) in porcine edible tissues pose a potential risk for human health. In this study, urine was selected as target matrix for routine residue monitoring of FF in pig, and a thin layer chromatography (TLC)-high-performance liquid chromatography (HPLC) method was developed for simultaneously determining FF and florfenicol amine (FFA) in porcine urine. The urine samples were extracted with ethyl acetate under alkaline environment. The extracts were enriched through evaporation, purified by TLC and analysed by HPLC at 225 nm. A Waters Symmetry C18 column was used for the separation of the two analytes. The mobile phase was acetonitrile-phosphate buffer mixtures (33.3: 66.7, v/v), and was pumped at 0.6 mL/min. The TLC-HPLC method was well validated and successfully applied to residue depletion study. Good analytical specificity was confirmed by the lack of interfering peaks at the retention times of FF and FFA. The standard curves showed good linearity (FF: y = 143064x – 1045.3, r= 0.9999; FFA: y = 275826x + 1888.8, r= 0.9999) over the range of 0.0625–8 μg/mL. The precision ranged from 0.83% to 11.66% and 2.19% to 8.75% for intraday and interday determination, respectively. The corresponding accuracy ranged from −13.38% to 10.78% and −12.15% to 7.14%, respectively. The limits of quantification (LOQs) for FF and FFA were 0.125 μg/mL. The residue depletion study showed that the concentrations of FF and FFA in urine were higher than those in edible tissues at three time points. This method was reliable, simple and cost efficient, and could be used to monitor FF residues in porcine edible tissue without slaughtering animals. TLC showed excellent purification efficiency and is expected to solve matrix interferences in veterinary drug residue analysis.

METHOD VALIDATION OF RIFAMPICIN ANALYSIS IN HUMAN PLASMA AND ITS APPLICATION IN BIOEQUIVALENCE STUDY

Objective: This research aims to validate the method for rifampicin analysis in plasma by using High-Performance Liquid Chromatography (HPLC) that can be used to study the bioequivalence of a generic tablet of rifampicin 450 mg “X” marketed in Indonesia.&#x0D; Methods: Bioequivalence test was analysed using HPLC equipped with UV-Vis detector at 377 nm. The mobile phase used was acetonitrile-phosphate buffer pH 6.8 (45:55) delivered at a flow rate of 1.5 ml/min. Bioequivalence test was conducted on a limited number of subjects (n=8). The subjects were divided into two groups randomly. The pharmacokinetic profiles of the test tablet and reference tablet were statistically calculated using SPSS program to see the test tablet and reference tablet were bioequivalence or not.&#x0D; Results: The developed HPLC method for rifampicin analysis in plasma was sufficiently valid based on the International Conference on Harmonization (ICH) and European Medicines Agency (EMA) guideline, with precision and accuracy values were % Relative Standard Deviation (% RSD = 1.40–13.04) and % Recovery (86.24–102.13), respectively. Meanwhile, the method was linear over studied concentration (0.05 to 10.26 µg/ml) with a coefficient of determination (R2) = 0.9984. The method also had good stability and sensitivity. The result of statistical calculation showed that the generic rifampicin tablet X was bioequivalence toward the reference tablet Rimactan 450 mg.&#x0D; Conclusion: The test rifampicin tablet that was, the generic tablet “X” was bioequivalence toward the reference rifampicin tablet “Rimactan”.

VALIDATION AND QUANTITATIVE ANALYSIS OF CARMINE AND RHODAMINE B IN LIPSTICK FORMULATION

Objective: The objective of this study was to validate reversed phase-high performance liquid chromatography (RP-HPLC) method using photo-diode array detector for the simultaneous determination of carmine (CAR) and rhodamine B (RHO B) in lipstick products.&#x0D; Methods: CAR and RHO B were analysed using Cosmosil C18 column (250 mm x 4.6 mm i.d., 5 µm) using Shimadzu LC 20AD chromatograph equipped with photo-diode array (PDA) detector at 245-600 nm. The mobile phase used was acetonitrile-phosphate buffer pH 3.4 (5.5:4.5 v/v) delivered isocratically at flow rate of 1.0 ml/min, using column temperature at 35 °C. The optimized HPLC condition was validated by determining several characteristics including selectivity, linearity, sensitivity expressed by the limit of detection and limit of quantification, precision and accuracy.&#x0D; Results: The optimized HPLC condition could separate CAR and RHO B successfully with resolution value&gt;2.0. The method was linear over studied concentration with a coefficient of determination (R2) 0.9999. The relative standard deviation (RSD) values were lower than those required RSD Horwitz and the recovery values were also acceptable, therefore, the developed method was precise and accurate.&#x0D; Conclusion: HPLC with PDA detector has been accurately and successfully used for quantitative analysis of CAR and RHO B. The developed method is simple and can be used for routine analysis of CAR and RHO B for quality assurance purposes.

Comparative analysis of stability of tricyclic analogues of acyclovir in an acidic environment

In the presented studies, the effect of hydrogen ions on the stability of the tricyclic aciclovir derivative (i.e. 3,9-dihydro-3-[(2-hydroxyethoxy)methyl]-6-(4-methoxyphenyl)-9-oxo-5H-imidazo[1,2-a]purine; 6-(4-MeOPh)-TACV) and its esters (acetyl (Ac-), iso-butyryl (iBut-), pivaloyl (Piv-), ethoxycarbonyl (Etc-) and nicotinoyl (Nic-)) has been assessed. The HPLC chromatographic method (Lichrospher RP-18 column, 5 μm, 250 mm × 4 mm) with UV detection (262 nm) was used to observe changes in the concentration of the tested compounds. The mobile phase consisted of acetonitrile–phosphate buffer (pH 6; 20 mM; 17 mM KH2PO4, 3 mM K2HPO4) (35:65, v/v). The studies were carried out in the pH range 0.42–1.38 in the water-organic environment at constant ionic strength (0.50 M). The observed rate constants of the degradation reactions of the tested compounds were determined, kinetic equations describing the dependence of kpH as a function of pH for the proper acid catalysis were determined and the parameters of the equations describing the reaction of formation the observed product were determined. The dependence of durability on the chemical structure of the tested compounds was indicated and the mechanism of observed reactions was proposed.